The focus of this project is on IgE-binding protein (epsilon-BP) with emphasis on its modulatory role on mast cell functions. Epsilon-BP is a 31,000 Mr protein identified in rat basophilic leukemia (RBL) cells and is now known as soluble lectin designated with different names by other laboratories and appears to have multiple functions. The protein has wide tissue distribution, is found on the cell surface and also secreted under certain conditions. Epsilon-BP has specificity for distinct oligosaccharide structures that have a terminal galactose not masked by sialic acids and it functions at least bivalently. In addition to binding IgE, epsilon-BP binds to surfaces of various cell types, including mast cells. First, the molecular basis for multivalency of epsilon-BP will be elucidated. We have evidence that epsilon-BP has a tendency to self-associate through intermolecular interactions involving the amino-- terminal domain, resulting in dimers or oligomers. This property of epsilon-BP will be further investigated, including equilibrium ultracentrifugation analysis and binding to lactosyl-Sepharose 4B containing varying densities of lactose. Another explanation for bivalency of epsilon-BP is the formation of a dimer through an inter-molecular disulfide linkage. The molecular mechanism for the covalent dimerization of epsilon-BP will be investigated. Second, cell surface glycoprotein ligands for epsilon-BP will be identified. Preliminary studies indicate that epsilon-BP is attached to glycoconjugates on the surface of mast cells and only a small number of different glycoproteins species on RBL cell surface are recognized by epsilon-BP. Significantly, one of these glycoproteins is the high affinity IgE receptor (Fc-epsilon-RI). We will focus on isolation and characterization of another epsilon-BP-reactive Mr 150,00 glycoprotein. We will then investigate possible variation in epsilon-BP recognition of FC-epsilon-RI on mast cells at various differentiation stages or activation status. Third, the function of epsilon-BP as a multifunctional modulator molecule will be established. Having demonstrated epsilon-BP's multivalent property and recognition of cell surface glycoproteins, especially Fc-epsilon-RI, we propose that this lectin has the potential to modulate cellular functions of mast cells mediated by these glycoproteins. We have already shown that epsilon-BP potentiates Fc-epsilon-RI-mediated mast cell activation. We will further substantiate this modulator role of epsilon-BP and investigate the mechanism for the observed potentiating effect.